PROJECT SUMMARY Costly and invasive corrective surgery is the only treatment option for pediatric maxillary bone deficiency (MBD), which occurs when there is restriction of upper jaw bone growth, or traumatic injury to the maxillary bone. Autogenous bone graft (ABG) harvest, commonly from the iliac crest, is the preferred surgical approach to replace or augment maxillary bone in children. While effective, ABG is associated with prolonged operative time, overnight stay in the hospital, severe hip pain, and risk of hip fracture during recovery. Our group developed a murine model of MBD by targeted deletion of Jagged1, a cell surface signaling ligand member of the Notch family, from the maxillary progenitor cells (maxillary mesenchymal pluripotent cells (MMPC)) using Wnt1-Cre; Jagged1 F/F mice. JAGGED1 mutations in humans are associated with MBD as well as pediatric bony pathology and increased incidence of bony fracture, demonstrating the importance of JAGGED1 signaling during bone formation and maintenance. Our long-term goal is to develop a regenerative, non- surgical approach to treat MBD. The overall objective in this application is to determine the mechanism of Jagged1 signaling to stimulate osteoblast commitment and develop a regenerative approach for in vivo delivery of Jagged1 therapy to induce local bone growth. The central hypothesis is that Jagged1 signaling terminally differentiates progenitor cells to form bone, and that targeted delivery of Jagged1-based therapy in a hydrogel will induce localized bone growth. The rationale for the proposed research is that a mechanistic determination of how Jagged1 stimulates osteoblast commitment will inform the development of bone tissue engineering approaches focused on Jagged1 delivery strategies to regenerate maxillary bone without ABG. Guided by strong preliminary data, the hypothesis will be tested by pursuing three specific aims: 1) Determine the mechanism of Jagged1-mediated MMPC osteoblast commitment; 2) Develop and test hydrogel-based Jagged1 osteoinductive scaffold in vitro and in vivo; 3) Characterize in vitro and in vivo ability of MMPC/MSC cell therapy to regenerate maxillary bone formation using PEG-MAL hydrogel delivery. In Aim 1 we will determine the mechanism and redundancy of Jagged1 signaling through the Notch receptors, intracellular osteogenic gene pathways and down stream Notch targets. In Aim 2, we will develop and test the in vitro and in vivo delivery of Jagged1-PEG-MAL to induce bone formation and in Aim 3, characterize the in vitro and in vivo ability of MMPC/MSC to form maxillary bone using PEG-MAL hydrogel delivery in a gene correction model. The proposed research is innovative by mechanistically describing the role of Jagged1 signaling during osteoblast commitment and developing Jagged1-based regenerative approaches to maxillary bone disease. The proposed research is significant because it is expected to be a progression of research that will develop potential therapies for pediatric maxillary bone deficiency, and may be relevant to all bone density and healing.